Partially protected peptide segments are often required as intermediates for the preparation of larger peptides or small proteins by segment condensation approaches, either in solution or on a polymeric support. F.M. Finn and K. Hofmann, In: The Proteins, 3rd Ed., Vol. 2., H. Neurath and R.L. Hills (eds.), Academic Press, N.Y. pp. 105-253 (1976); G. Barany et al., Int. J. Peptide Protein Res., 30:705-739 (1987) Solid-phase synthesis is widely acknowledged to offer the best prospects for rapid and efficient assembly of peptide chains, but until relatively recently, the needed levels of selectivity in conditions for deprotection of the N.sup..alpha. -amino group and side-chains of amino acids, and in procedures for detachment of peptides from the support, have not been available. Such conditions require that a peptide can be cleaved successfully to furnish a free C.sup..alpha. -carboxyl group with all other functional groups remaining protected, and that the resultant intermediate can be purified before its further use in segment condensation. T. Kubiak et al., Biochemistry, 26:7849-7855 (1987); N. Kneib-Cordonier et al., Int. J. Peptide Protein Res., 35:527-538 (1990); G.B. Field and R.L. Noble, Int. J. Peptide Protein Res., 35:161-214 (1990).
In recent years, a strategy employing the orthogonal combination of base-labile 9-fluorenylmethyloxycarbonyl (Fmoc) for N.sup..alpha. -amino protection and acid-labile tert-butyl (tBu) derivatives for side-chain protection has been used. An "orthogonal" system is defined as one using two or more independent classes of protecting groups that are removed by different chemical mechanisms. This combination avoids the relatively harsh final cleavage conditions of the more conventional strategy based on the graduated lability to acid of tert-butyloxycarbonyl (Boc) for N.sup..alpha. -amino protection and benzyl or cyclohexyl derivatives for side-chain protection. In the Fmoc/tBu strategy, a third dimension of orthogonality can be provided by use of ortho-nitrobenzyl (photolabile), silicon-containing (fluoride-labile), or allyl-derived (cleaved with Pd(O)) anchoring linkages. R. Ramage et al., Tetrahedron Lett., 28:4105-4108 (1987), D.G. Mullen and G. Barany, J. Org. Chem., 53:5240-5248 (1988); H. Kunz and B. Dombo, Agnew. Chem. Int. Ed. Engl., 27:711-713 (1988); B. Blankemeyer-Menge and R. Frank, Tetrahedron Lett., 28:5871-5874 (1988). However, the aforementioned orthogonally cleavable anchors are prepared by multi-step routes, and in some cases, cannot be satisfactorily applied to solid-phase synthesis of protected peptide segments.
An alternative approach utilizes anchoring linkages, or handles, that are cleaved with extremely dilute acid. This strategy requires exquisite "fine-tuning" of the anchor structure and the corresponding removal conditions. For example, a (4-hydroxymethyl-3-methoxyphenoxy)acetic acid handle and the closely related 2-methoxy-4-alkoxybenzyl alcohol (SASRIN) support have been used and lead to bis(alkoxy)benzyl ester anchoring linkages which cleave with 1% (v/v) trifluoracetic acid (TFA) in dichloromethane. E. Atherton et al., In: Solid Phase Peptide Synthesis, IRL Press Oxford, U.K. (1989); R.C. Sheppard and B.J. Williams J. Chem. Soc. Chem. Commun., pp. 587-589 (1982); M. Mergler et al., Tetrahedron Lett., 29:4009-4012 (1988). These cleavage conditions promote premature side-chain deprotection at Lys(Boc) and Tyr(tBu), however. R.C. Sheppard and B.J. Williams, J. Chem. Soc. Chem. Commun., ibid. Other systems using a trialkoxydiphenylmethyl ester handle or an ortho-chlorotrityl handle lead to esters which cleave with 10% (v/v) acetic acid in dichloromethane, but also cleave prematurely in the presence of a free C.sup..alpha. -carboxyl group of an incoming protected amino acid during each coupling step. H. Rink, Tetrahedron Lett., 28:3787-3790 (1987); K. Barlos et al., Tetrahedron Lett., 80:3943-3946 (1989). A handle which permits efficient cleavage of a peptide from the solid support, without causing deprotection of side-chain groups or N.sup..alpha. terminal groups, would be valuable for preparing protected peptide intermediates and for other applications where deprotection is not desired.